Abstract: In the United States (U.S.), transgender women (TW) are one of the populations most affected by HIV infection. The high prevalence of HIV infection among U.S. TW is driven by, and/or concomitant with, structural barriers that limit access to HIV prevention, care, and health services. Despite an emergence of research to characterize the HIV epidemic among TW, the majority of studies are cross-sectional designs and typically include small sample sizes, often subsuming TW among broader risk groups, such as men who have sex with men. These practices have thwarted identification of acceptable, effective recruitment and study methods for use among TW and prevented temporal assessment, causal inference, and generalizability of study findings to the TW population. To date, there is no robust estimate of HIV incidence and no intervention with evidence of efficacy for the prevention of HIV acquisition among TW. The proposed American Cohort study will address these limitations by establishing a multi-site, longitudinal cohort of TW spanning eastern and southern U.S. (Boston, New York City, Baltimore-Washington, Atlanta, and Miami metropolitan areas) to characterize HIV incidence and risk factors for HIV acquisition, access to biobehavioral HIV prevention methods, and linkage to care for those who HIV seroconvert. The cohort will include a racially/ethnically and culturally diverse sample of TW, supported by the use of technology-infused recruitment and retention methods. The specific aims are: 1) To determine the efficiency and acceptability of novel, technology-infused recruitment methods to enroll HIV- uninfected TW into a prospective cohort. 2) To describe the demographic, socioeconomic, behavioral, and physical and mental health profiles of HIV-uninfected TW in the first, multi-site cohort of TW in the eastern and southern U.S. 3) To estimate HIV incidence among TW in high-risk eastern and southern U.S. areas, trends in incidence, and associated individual, social, and structural risk factors. 4) To estimate the HIV Prevention Continuum among HIV-uninfected participants, and the HIV Care Continuum among newly HIV-infected TW. To achieve these aims we will recruit 1,750 TW who will be enrolled in either a HIV-uninfected cohort or an HIV-infected cross-sectional comparison group. These data will be used to assess differences between cohort participants and the wider population and for cross-sectional incidence estimation at enrollment. The HIV- uninfected cohort will be followed for at least 24 months to estimate HIV incidence, trends, and risk factors for HIV acquisition. Cohort participants who seroconvert will be followed for an additional 6 months to assess prospective engagement in the HIV Care Continuum. Study findings will provide critical epidemiologic parameters for future HIV prevention research among TW, provide a platform upon which other research questions can be explored, and inform the development of evidence-based and acceptable HIV interventions to reduce HIV acquisition among TW in the U.S.